VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

CBV016Membrane potential plays a fundamental role inregulation and maintenance of bacterial morphologyH. Strahl*, L. HamoenCenter for Bacterial Cell Biology, Newcastle University, Newcastle uponTyne, United KingdomThe emerging knowledge about the distinct localisation of proteins and othercellular components has radically changed our view of bacterial cells. Theorganisation of different cellular functions to specific areas of the cellreflects the existence of a well-defined cellular architecture. However, thepresence of a high level of organisation is fundamentally linked to theenergy required for its maintenance. In addition, many cellular structures aredynamic in their localisation and macromolecular structure, furtheremphasizing the critical role of energy supply. The role of high energyphosphates like ATP and GTP in maintaining the cell architecture has beenpreviously analysed in great detail. However, all living organisms alsoutilise another fundamental energy source, the transmembrane proton motiveforce (pmf). This second major cellular energy source is crucial for variousprocesses including transport, signalling and ATP-synthesis across alldomains of life. Although cell membranes and membrane proteins play acentral role in bacterial morphology, nothing is known about the role of pmfin these processes. A detailed analysis of key morphological proteins inBacillus subtilis revealed a drastic effect on their localisation when the pmfwas dissipated. Based on these results, we propose a novel function of themembrane potential in regulation and maintenance of bacterial morphology.Strahl H, Hamoen LW. (2010) Membrane potential is important for bacterialcell division. PNAS 107:12281-12286.CBV017Vip1-like 1/3 inositol polyphosphate kinases regulate thedimorphic switch in yeastsJ. Pöhlmann*, U. FleigInstitute of Functional Genomics of Microorganisms, Heinrich-Heine-University, Düsseldorf, GermanyIt has long been known that the environmentally induced transition of fungalgrowth forms is an essential initial requirement for pathogenesis. The abilityto undergo a dramatic morphological change from a single cell form to amulti-cellular invasive form in response to extrinsic cues is conserved infungi and also found in non-pathogenic model yeasts such as S. pombe andS. cerevisiae. Here we describe the identification and characterization of theS. pombe Asp1 protein as a key regulator of the dimorphic switch. Asp1 is amember of the highly conserved Vip1 family of 1/3 inositol polyphosphatekinases, which generate specific inositol pyrophosphates that have beenshown to regulate cyclin-CDK complexes. Vip1-like proteins have a dualdomain structure consisting of an N-terminal „rimK”/ATP-graspsuperfamily domain and a C-terminal part with homology to histidine acidphosphatases.Asp1, which acts downstream of the cAMP PKA pathway, isessential for the transition to the pseudohyphal invasive growth mode undernutrient limitation. Intriguingly, an increase in the cellular amounts of Asp1generated inositol pyrophosphates increases the cellular response thusimplying that these molecules might act as second messengers. Remarkablythe Asp1 kinase activity is regulated negatively by its C-terminal domain.Thus the fine tuning of the cellular response to environmental cues ismodulated by the same protein. Interestingly, the S. cerevisiae Vip1 familymember is also required for the dimorphic switch in this yeast. Therefore wepropose Vip1 family members have a general role in regulating fungaldimorphism and are presently testing this in a number of fungi.CBP001Coordinated separation - the late stage of bacterial celldivisionA. Möll* 1,2 , S. Schlimpert 1,2 , A. Briegel 3 , G.J. Jensen 3 , M. Thanbichler 1,21 Department of Biology, Philipps-University, Marburg, Germany2 Research Group Prokaryotic Cell Biology, Max Planck Institute forTerrestrial Microbiology, Marburg, Germany3 Division of Biology and California Institute of Technology, Howard HughesMedical Institute, Pasadena, USAIn the late stages of bacterial cell division, the remodelling of the cell wallrequires a delicate balance between synthesis and degradation ofpeptidoglycan. Only few components of the protein network orchestratingthis process have been identified, and the mode of their spatial and temporalregulation remains unclear. To address this issue, we investigate the functionof cell division proteins in the gram-negative model organism Caulobactercrescentus.Cell wall peptidoglycan is a structural element preserving cell integrity andcontributing to cell shape. Additionally, it serves as a scaffold for anchoringproteins that are part of the cell envelope. To identify factors involved in thelate stage of cell division, we focused on proteins containing predictedpeptidoglycan-binding domains. Using fluorescence microscopy, weselected promising candidates that localized to midcell during cell divisionand subsequently examined them in more detail.Based on this approach, we identified and characterized a structuralhomologue of the late essential cell division protein FtsN from Escherichiacoli in C. crescentus. FtsN was previously thought to be poorly conservedoutside the enteric bacteria. However, a database search based on the typicalstructural features shared by E. coli and C. crescentus FtsN showed thatFtsN-like proteins are in fact widespread among all proteobacteria [1].Building on these results, we identified an interaction partner of FtsN,named DipM, for division- and polarity-related metallopeptidase. DipMrequires FtsN for midcell localization. Interestingly, in the absence of DipM,invagination of the cell wall and outer membrane is delayed, leading tosevere division and polarity defects [2]. These results provide more evidencefor a key role of FtsN in the regulation of cell wall remodelling during thefinal stage of cell division.[1] Möll, A., and M. Thanbichler (2009): FtsN-like proteins are conserved components of the celldivision machinery in proteobacteria. Mol Microbiol 72: 1037-1053.[2] Möll, A. et al (2010): DipM, a new factor required for peptidoglycan remodelling during celldivision in Caulobacter crescentus. Mol Microbiol 77: 90-107.CBP002Mechanism of Gradient Formation by the CaulobacterCell Division Inhibitor MipZD. Kiekebusch* 1 , K.A. Michie 2 , L.-O. Essen 3 , J. Löwe 2 , M. Thanbichler 11 Max Planck Institute for Terrestrial Microbiology and Laboratory forMicrobiology, Philipps-University Marburg, Marburg, Germany2 Medical Research Council, Cambridge, United Kingdom3 Department of Chemistry, Structural Biochemistry, Philipps UniversityMarburg, Marburg, GermanyIntracellular protein gradients play a critical role in the spatial organizationof both prokaryotic and eukaryotic cells, but in many cases the mechanismsunderlying their formation are still unclear. Recently, a bipolar gradient ofthe Walker ATPase MipZ was found to be required for proper division siteplacement in the differentiating bacterium Caulobacter crescentus. MipZinteracts with a kinetochore-like nucleoprotein complex formed by the DNApartitioning protein ParB in proximity of the chromosomal origin ofreplication. Upon entry into S-phase, the two newly duplicated originregions are partitioned and sequestered to opposite cell poles, giving rise to abipolar distribution of MipZ with a defined concentration minimum at thecell center. Acting as a direct inhibitor of divisome formation, MipZ thuseffectively confines cytokinesis to the midcell region. Building on thecrystal structures of the apo and ATP-bound protein, we have dissected therole of nucleotide binding and hydrolysis in MipZ function. Our findingsindicate that gradient formation results from alternation of MipZ between amonomeric and dimeric form that display marked differences in theirinteraction networks and diffusion rates. As a consequence, MipZ undergoesan elaborate localization cycle, involving its oscillation between the polarParB complexes and pole-distal regions of the nucleoid. The MipZ gradientthus represents the steady-state distribution of molecules in a highlydynamic system, providing a general mechanism for the establishment ofprotein gradients within the confined space of the bacterial cytoplasm.CBP003Functional analysis of SPFH domain-containing proteins,Flotillin and Stomatin, in Aspergillus nidulansN. Takeshita*, R. FischerDeparment of Microbiology, Karlsruhe Institute of Technology (KIT),Karlsruhe, GermanyPolarized growth of filamentous fungi depends on the microtubule and theactin cytoskeleton along with their associated motor proteins. Apicalmembrane-associated landmark proteins, so-called „cell end markers” linkthe two cytoskeletons. Our latest results indicate that apical sterol-richmembrane domains (SRDs) play important roles in polarized growth andspektrum | Tagungsband 2011